*Target (protein/gene name): alkaline phosphatase *NCBI Gene # or RefSeq#: Gene ID: 248 *Protein ID (NP or XP #) or Wolbachia#: *Organism (including strain): Clostridium botulinum Etiologic Risk Group (see link below): 2 */Disease Information (sort of like the Intro to your Mini Research Write up):
Botulism is a disease caused by the bacteria clostridium botulinum. Botulism occurs in humans when foods contaminated by this bacterium is consumed. The bacteria attack the immune system by releasing toxic spores in the intestines. Different types of diseases caused by this bacteria include foodborne botulism, infact botulism, wound botulism, etc. Symptoms of these diseases involve dropping eyelids, nausea and vomiting, muscle weakness, etc. The main treatment used to treat botulism is antibiotics.
*EC#: 3.1.3.1. Link to BRENDA EC# page: -- Show screenshot of BRENDA enzyme mechanism schematic Enzyme Assay information (spectrophotometric, coupled assay ?, reagents): -- link to Sigma (or other company) page for assay (see Sigma links below) http://www.sigmaaldrich.com/technical-documents/protocols/biology/enzymatic-assay-of-alkaline-phosphatase-diethanolamine-assay.html -- -or link (or citation) to paper that contains assay information -- links to assay reagents (substrates) pages. --- List cost and quantity of substrate reagents, supplier, and catalog #
Structure (PDB or Homology model) -- PDB # or closest PDB entry if using homology model: -- For Homology Model option: ---- Show pairwise alignment of your BLASTP search in NCBI against the PDB ---- Query Coverage: ---- Max % Identities: ---- % Positives ---- Chain used for homology:
Current Inhibitors: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1162816/pdf/biochemj00403-0154.pdf Expression Information (has it been expressed in bacterial cells): Purification Method: Image of protein (PyMol with features delineated and shown separately): *Amino Acid Sequence (paste as text only - not as screenshot or as 'code'): >5TJ3:A|PDBID|CHAIN|SEQUENCE MDIGIDSDPQKTNAVPRPKLVVGLVVDQMRWDYLYRYYSKYGEGGFKRMLNTGYSLNNVHIDYVPTVTAIGHTSIFTGSV PSIHGIAGNDWYDKELGKSVYCTSDETVQPVGTTSNSVGQHSPRNLWSTTVTDQLGLATNFTSKVVGVSLKDRASILPAG HNPTGAFWFDDTTGKFITSTYYTKELPKWVNDFNNKNVPAQLVANGWNTLLPINQYTESSEDNVEWEGLLGSKKTPTFPY TDLAKDYEAKKGLIRTTPFGNTLTLQMADAAIDGNQMGVDDITDFLTVNLASTDYVGHNFGPNSIEVEDTYLRLDRDLAD FFNNLDKKVGKGNYLVFLSADHGAAHSVGFMQAHKMPTGFFVEDMKKEMNAKLKQKFGADNIIAAAMNYQVYFDRKVLAD SKLELDDVRDYVMTELKKEPSVLYVLSTDEIWESSIPEPIKSRVINGYNWKRSGDIQIISKDGYLSAYSKKGTTHSVWNS YDSHIPLLFMGWGIKQGESNQPYHMTDIAPTVSSLLKIQFPSGAVGKPITEVIGRIEGRSAWSHPQFEK
*length of your protein in Amino Acids: 549 Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 61254.95 Molar Extinction coefficient of your protein at 280 nm wavelength: 104740 TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it. *CDS Gene Sequence (paste as text only):
*GC% Content for gene: *CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): *GC% Content for gene (codon optimized):
Do Not Need this info for Spring (but still copy these lines to your Target page for now) Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers): (link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol) -- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences): **
*NCBI Gene # or RefSeq#: Gene ID: 248
*Protein ID (NP or XP #) or Wolbachia#:
*Organism (including strain): Clostridium botulinum
Etiologic Risk Group (see link below): 2
*/Disease Information (sort of like the Intro to your Mini Research Write up):
Botulism is a disease caused by the bacteria clostridium botulinum. Botulism occurs in humans when foods contaminated by this bacterium is consumed. The bacteria attack the immune system by releasing toxic spores in the intestines. Different types of diseases caused by this bacteria include foodborne botulism, infact botulism, wound botulism, etc. Symptoms of these diseases involve dropping eyelids, nausea and vomiting, muscle weakness, etc. The main treatment used to treat botulism is antibiotics.
Link to TDR Targets page (if present): http://www.tdrtargets.org/targets/view?gene_id=285537
Link to Gene Database page (NCBI, EuPath databases -e.g. TryTryp, PlasmoDB, etc - or PATRIC, etc.)
Essentiality of this protein:
Is it a monomer or multimer as biological unit? (make prediction at http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html): Monomer
Complex of proteins?:
Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism): https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1162816/pdf/biochemj00403-0154.pdf
*EC#: 3.1.3.1.
Link to BRENDA EC# page:
-- Show screenshot of BRENDA enzyme mechanism schematic
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
-- link to Sigma (or other company) page for assay (see Sigma links below)
http://www.sigmaaldrich.com/technical-documents/protocols/biology/enzymatic-assay-of-alkaline-phosphatase-diethanolamine-assay.html
-- -or link (or citation) to paper that contains assay information
-- links to assay reagents (substrates) pages.
--- List cost and quantity of substrate reagents, supplier, and catalog #
Structure (PDB or Homology model)
-- PDB # or closest PDB entry if using homology model:
-- For Homology Model option:
---- Show pairwise alignment of your BLASTP search in NCBI against the PDB
---- Query Coverage:
---- Max % Identities:
---- % Positives
---- Chain used for homology:
http://www.rcsb.org/pdb/explore/explore.do?structureId=5TJ3
Current Inhibitors:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1162816/pdf/biochemj00403-0154.pdf
Expression Information (has it been expressed in bacterial cells):
Purification Method:
Image of protein (PyMol with features delineated and shown separately):
*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
>5TJ3:A|PDBID|CHAIN|SEQUENCE
MDIGIDSDPQKTNAVPRPKLVVGLVVDQMRWDYLYRYYSKYGEGGFKRMLNTGYSLNNVHIDYVPTVTAIGHTSIFTGSV
PSIHGIAGNDWYDKELGKSVYCTSDETVQPVGTTSNSVGQHSPRNLWSTTVTDQLGLATNFTSKVVGVSLKDRASILPAG
HNPTGAFWFDDTTGKFITSTYYTKELPKWVNDFNNKNVPAQLVANGWNTLLPINQYTESSEDNVEWEGLLGSKKTPTFPY
TDLAKDYEAKKGLIRTTPFGNTLTLQMADAAIDGNQMGVDDITDFLTVNLASTDYVGHNFGPNSIEVEDTYLRLDRDLAD
FFNNLDKKVGKGNYLVFLSADHGAAHSVGFMQAHKMPTGFFVEDMKKEMNAKLKQKFGADNIIAAAMNYQVYFDRKVLAD
SKLELDDVRDYVMTELKKEPSVLYVLSTDEIWESSIPEPIKSRVINGYNWKRSGDIQIISKDGYLSAYSKKGTTHSVWNS
YDSHIPLLFMGWGIKQGESNQPYHMTDIAPTVSSLLKIQFPSGAVGKPITEVIGRIEGRSAWSHPQFEK
*length of your protein in Amino Acids: 549
Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 61254.95
Molar Extinction coefficient of your protein at 280 nm wavelength: 104740
TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.
*CDS Gene Sequence (paste as text only):
*GC% Content for gene:
*CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only):
*GC% Content for gene (codon optimized):
Do Not Need this info for Spring (but still copy these lines to your Target page for now)
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)
-- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):
**